Exploiting Differential Signal Filtering (DSF) and Image Structure Filtering (ISF) Methods for Untargeted Mass Spectrometry Imaging of Bacterial Metabolites.

Matrix-assisted laser desorption/ionization (MALDI) mass spectrometry imaging (MSI) is a label-free technique, producing images where pixels contain mass spectra. The technique allows the visualization of the spatial distribution of (bio)molecules from metabolites to proteins, on surfaces such as tissues sections or bacteria culture media. One particularly exciting example of MALDI-MSI use rests on its potential to localize ionized compounds produced during microbial interactions and chemical communication, offering a molecular snapshot of metabolomes at a given time. The huge size and the complexity of generated MSI data make the processing of the data challenging, which requires the use of computational methods. Despite recent advances, currently available commercial software relies mainly on statistical tools to identify patterns, similarities, and differences within data sets. However, grouping m/z values unique to a given data set according to microbiological contexts, such as coculture experiments, still requires tedious manual analysis. Here we propose a nontargeted method exploiting the differential signals between negative controls and tested experimental conditions, i.e., differential signal filtering (DSF), and a scoring of the ion images using image structure filtering (ISF) coupled with a fold change score between the controls and the conditions of interest. These methods were first applied to coculture experiments involving Escherichia coli and Streptomyces coelicolor, revealing specific MS signals during bacterial interaction. Two case studies were also investigated: (i) cellobiose-mediated induction for the pathogenicity of Streptomyces scabiei, the causative agent of common scab on root and tuber crops, and (ii) iron-repressed production of siderophores of S. scabiei. This report proposes guidelines for MALDI-MSI data treatment applied in the case of microbiology contexts, with enhanced ion peak annotation in specific culture conditions. The strengths and weaknesses of the methods are discussed.

Nitrogen sources enhance siderophore-mediated competition for iron between potato common scab and late blight causative agents.

How do pathogens affecting the same host interact with each other? We evaluated here the types of microbe-microbe interactions taking place between Streptomyces scabiei and Phytophthora infestans, the causative agents of common scab and late blight diseases in potato crops, respectively. Under most laboratory culture conditions tested, S. scabiei impaired or completely inhibited the growth of P. infestans by producing either soluble and/or volatile compounds. Increasing peptone levels correlated with increased inhibition of P. infestans. Comparative metabolomics showed that production of S. scabiei siderophores (desferrioxamines, pyochelin, scabichelin, and turgichelin) increased with the quantity of peptone, thereby suggesting that they participate in the inhibition of the oomycete growth. Mass spectrometry imaging further uncovered that the zones of secreted siderophores and of P. infestans growth inhibition coincided. Moreover, either the repression of siderophore production or the neutralization of their iron-chelating activity led to a resumption of P. infestans growth. Replacement of peptone by natural nitrogen sources such as ammonium nitrate, sodium nitrate, ammonium sulfate, and urea also triggered siderophore production in S. scabiei. Interestingly, nitrogen source-induced siderophore production also inhibited the growth of Alternaria solani, the causative agent of the potato early blight. Overall, our work further emphasizes the importance of competition for iron between microorganisms that colonize the same niche. As common scab never alters the vegetative propagation of tubers, we propose that S. scabiei, under certain conditions, could play a protective role for its hosts against much more destructive pathogens through exploitative iron competition and volatile compound production.

Lipopeptides as rhizosphere public goods for microbial cooperation.

IMPORTANCE: Here, we provide new insights into the possible fate of cyclic lipopeptides as prominent specialized metabolites from beneficial bacilli and pseudomonads once released in the soil. Our data illustrate how the B. velezensis lipopeptidome may be enzymatically remodeled by Streptomyces as important members of the soil bacterial community. The enzymatic arsenal of S. venezuelae enables an unsuspected extensive degradation of these compounds, allowing the bacterium to feed on these exogenous products via a mechanism going beyond linearization, which was previously reported as a detoxification strategy. As soils are carbon-rich and nitrogen-poor environments, we propose a new role for cyclic lipopeptides in interspecies interactions, which is to fuel the nitrogen metabolism of a part of the rhizosphere microbial community. Streptomyces and other actinomycetes, producing numerous peptidases and displaying several traits of beneficial bacteria, should be at the front line to directly benefit from these metabolites as "public goods" for microbial cooperation.

Exploring the Phytobeneficial and Biocontrol Capacities of Endophytic Bacteria Isolated from Hybrid Vanilla Pods.

In this study, 58 endophytic bacterial strains were isolated from pods of two hybrid vanilla plants from Madagascar, Manitra ampotony and Tsy taitra. They were genetically characterized and divided into four distinct phylotypes. Three were associated to genus Bacillus species, and the fourth to the genus Curtobacterium. A selection of twelve strains corresponding to the identified genetic diversity were tested in vitro for four phytobeneficial capacities: phosphate solubilisation, free nitrogen fixation, and phytohormone and siderophore production. They were also evaluated in vitro for their ability to biocontrol the growth of the vanilla pathogenic fungi, Fusarium oxysporum f. sp. radicis vanillae and Cholletotrichum orchidophilum. Three bacteria of phylotype 4, m62a, m64 and m65, showed a high nitrogen fixation capacity in vitro, similar to the Pseudomonas florescens F113 bacterium used as a control (phospate solubilizing efficiency respectively 0.50 ± 0.07, 0.43 ± 0.07 and 0.40 ± 0.06 against 0.48 ± 0.03). Strain t2 related to B. subtilis showed a higher siderophore production than F113 (respectively 1.40 ± 0.1 AU and 1.2 ± 0.1 AU). The strain m72, associated with phylotype 2, showed the highest rate of production of Indole-3-acetic acid (IAA) in vitro. Bacteria belonging to the pylotype 4 showed the best capacity to inhibit fungal growth, especially the strains m62b m64 and t24, which also induced a significant zone of inhibition, suggesting that they may be good candidates for controlling fungal diseases of vanilla. This competence was highlighted with spectral imaging showing the production of lipopeptides (Iturin A2 and A3, C16 and C15-Fengycin A and C14 and C15-Surfactin) by the bacterial strains m65 confronted with the pathogenic fungi of vanilla.

An evaluation of the self-assembly enhancing properties of cell-derived hexameric amyloid-ß.

A key hallmark of Alzheimer's disease is the extracellular deposition of amyloid plaques composed primarily of the amyloidogenic amyloid-ß (Aß) peptide. The Aß peptide is a product of sequential cleavage of the Amyloid Precursor Protein, the first step of which gives rise to a C-terminal Fragment (C99). Cleavage of C99 by γ-secretase activity releases Aß of several lengths and the Aß42 isoform in particular has been identified as being neurotoxic. The misfolding of Aß leads to subsequent amyloid fibril formation by nucleated polymerisation. This requires an initial and critical nucleus for self-assembly. Here, we identify and characterise the composition and self-assembly properties of cell-derived hexameric Aß42 and show its assembly enhancing properties which are dependent on the Aß monomer availability. Identification of nucleating assemblies that contribute to self-assembly in this way may serve as therapeutic targets to prevent the formation of toxic oligomers.